Generation and transcriptomic characterization of MIR137 knockout miniature pig model for neurodevelopmental disorders.

BACKGROUND: Neurodevelopmental disorders (NDD), such as autism spectrum disorders (ASD) and intellectual disorders (ID), are highly debilitating childhood psychiatric conditions. Genetic factors are recognized as playing a major role in NDD, with a multitude of genes and genomic regions implicated. While the functional validation of NDD-associated genes has predominantly been carried out using mouse models, the significant differences in brain structure and gene function between mice and humans have limited the effectiveness of mouse models in exploring the underlying mechanisms of NDD. Therefore, it is important to establish alternative animal models that are more evolutionarily aligned with humans. RESULTS: In this study, we employed CRISPR/Cas9 and somatic cell nuclear transplantation technologies to successfully generate a knockout miniature pig model of the MIR137 gene, which encodes the neuropsychiatric disorder-associated microRNA miR-137. The homozygous knockout of MIR137 (MIR137-/-) effectively suppressed the expression of mature miR-137 and led to the birth of stillborn or short-lived piglets. Transcriptomic analysis revealed significant changes in genes associated with neurodevelopment and synaptic signaling in the brains of MIR137-/- miniature pig, mirroring findings from human ASD transcriptomic data. In comparison to miR-137-deficient mouse and human induced pluripotent stem cell (hiPSC)-derived neuron models, the miniature pig model exhibited more consistent changes in critical neuronal genes relevant to humans following the loss of miR-137. Furthermore, a comparative analysis identified differentially expressed genes associated with ASD and ID risk genes in both miniature pig and hiPSC-derived neurons. Notably, human-specific miR-137 targets, such as CAMK2A, known to be linked to cognitive impairments and NDD, exhibited dysregulation in MIR137-/- miniature pigs. These findings suggest that the loss of miR-137 in miniature pigs affects genes crucial for neurodevelopment, potentially contributing to the development of NDD. CONCLUSIONS: Our study highlights the impact of miR-137 loss on critical genes involved in neurodevelopment and related disorders in MIR137-/- miniature pigs. It establishes the miniature pig model as a valuable tool for investigating neurodevelopmental disorders, providing valuable insights for potential applications in human research.

Nuclear microRNA-mediated transcriptional control determines adult microglial homeostasis and brain function.

Microglia are versatile regulators in brain development and disorders. Emerging evidence links microRNA (miRNA)-mediated regulation to microglial function; however, the exact underlying mechanism remains largely unknown. Here, we uncover the enrichment of miR-137, a neuropsychiatric-disorder-associated miRNA, in the microglial nucleus, and reveal its unexpected nuclear functions in maintaining the microglial global transcriptomic state, phagocytosis, and inflammatory response. Mechanistically, microglial Mir137 deletion increases chromatin accessibility, which contains binding motifs for the microglial master transcription factor Pu.1. Through biochemical and bioinformatics analyses, we propose that miR-137 modulates Pu.1-mediated gene expression by suppressing Pu.1 binding to chromatin. Importantly, we find that increased Pu.1 binding upregulates the target gene Jdp2 (Jun dimerization protein 2) and that knockdown of Jdp2 significantly suppresses the impaired phagocytosis and pro-inflammatory response in Mir137 knockout microglia. Collectively, our study provides evidence supporting the notion that nuclear miR-137 acts as a transcriptional modulator and that this microglia-specific function is essential for maintaining normal adult brain function.

Motorized and non-motorized mixed traffic characteristics and lateral opening spacing calculation for the section of arterial highway through small towns.

A large number of motor vehicles and non-motorized vehicles mixed in the section of arterial highway crossing the town leads to many traffic problems. Therefore, it is necessary to set up a side divider between motorized and non-motorized lanes, and the appropriate spacing of lateral crossing openings to meet the needs of non-motorized vehicles crossing the highway has become a key issue that must be resolved. This paper investigates the traffic flow characteristics of mixed traffic flow on arterial highways through small town sections, and from the two dimensions of highway access efficiency and the psychological characteristics of cyclists, it calibrates the setting range of the spacing of non-motorized lateral crossing openings under different design speeds, which is used to regulate the behavior of non-motorized vehicles crossing the street, improve the safety level of the highway, reduce the lateral interference of the highway, and improve the road access efficiency. The accuracy of the research results is verified by microscopic simulation experiments, which proves that they can meet good expectations in practical engineering. The research results have theoretical significance and reference value for improving the status quo of machine-non-mixed traffic in the section of arterial highway passing through small towns, and enhancing the efficiency and safety of highway traffic. It also provides corresponding reference for the areas facing similar problems worldwide.

Metal-Free Phosphination and Continued Functionalization of Pyridine: A Theoretical Study.

This study investigates the mechanism of metal-free pyridine phosphination with P(OEt)3, PPh3, and PAr2CF3 using density functional theory calculations. The results show that the reaction mechanism and rate-determining step vary depending on the phosphine and additive used. For example, phosphination of pyridine with P(OEt)3 occurs in five stages, and ethyl abstraction is the rate-determining step. Meanwhile, 2-Ph-pyridine phosphination with PPh3 is a four-step reaction with proton abstraction as the rate-limiting step. Energy decomposition analysis of the transition states reveals that steric hindrance in the phosphine molecule plays a key role in the site-selective formation of the phosphonium salt. The mechanism of 2-Ph-pyridine phosphination with PAr2CF3 is similar to that with PPh3, and analyses of the effects of substituents show that electron-withdrawing groups decreased the nucleophilicity of the phosphine, whereas aryl electron-donating groups increased it. Finally, TfO- plays an important role in the C-H fluoroalkylation of pyridine, as it brings weak interactions.

Origin, Expansion, and Divergence of ETHYLENE-INSENSITIVE 3 (EIN3)/EIN3-LIKE Transcription Factors During Streptophytes Evolution.

The transition of plants to land required several regulatory adaptive mechanisms. Little is known about these mechanisms, but they no doubt involved the evolution of transcription factor (TF) families. ETHYLENE-INSENSITIVE 3 (EIN3)/EIN3-LIKE (EIL) transcription factors (TFs) are core components of the ethylene signaling pathway that play important roles in almost every aspect of plant development and environmental responses by regulating the transcription of numerous genes. However, the evolutionary history of EIN3/EIL TFs, which are present in a wide range of streptophytes, is still not clear. Here, to explore the evolution and functions of EIN3/EIL TFs, we performed phylogenetic analysis of these TFs and investigated their gene and protein structures as well as sequence features. Our results suggest that the EIN3/EIL TF family was already was already present in the ancestor of streptophyte algae. Phylogenetic analysis divided the EIN3/EIL TFs into three groups (Group A-C). Analysis of gene and protein structure revealed that most of the structural features of these TFs had already formed in ancient lineages. Further investigation suggested that all groups have undergone several duplication events related to whole-genome duplications in plants, generating multiple, functionally diverse gene copies. Therefore, as plants colonized terrestrial habitats and evolved key traits, the EIN3/EIL TF family expanded broadly via multiple duplication events, which could have promoted their fundamental neo- and sub-functionalization to help plants adapt to terrestrial life. Our findings shed light on the functional evolution of the EIN3/EIL TF family in the streptophytes.